title: Cigarette smoke induces oxidative stress and apoptosis in normal term fetal membranes.
abstract: Cigarette smoking and bacterial infections are two major risk factors associated with preterm prelabor rupture of membranes (pPROM). We hypothesized that exposure of fetal membranes to cigarette smoke extracts might induce oxidative stress (OS) and fetal membrane apoptosis, culminating in an alternate pathway to that commonly activated by infection. To test this, we characterized the production of prostanoids and biomarkers of apoptosis in normal term human fetal membrane explant cultures. Fetal membrane explants collected at term (from cesarean deliveries, not in labor) were stimulated with cigarette smoke extract (CSE) for 24 h. Two classes of prostanoids, F2-Isoprostane (F2-IsoP), a marker of OS and PGF2α, a classical uterotonin, were measured by gas chromatography/mass spectrometry. Western blot analyses of tissue lysates were performed to quantify the anti-apoptotic protein Bcl2 and actin (as a control). Fetal membrane apoptosis was detected by immunohistochemistry for active caspase 3 and confirmed by TUNEL staining for nuclear fragmentation. CSE exposure resulted in significantly more F2-IsoP production from fetal membranes (242.8 ± 79.3 pg/ml/mg of total membrane protein) compared to unstimulated controls (131.5 ± 53.1 pg/ml/mg; p < 0.0001). By contrast, PGF2α was not different in CSE vs. controls (1083 ± 527 vs. 1136 ± 835 pg/ml/mg of protein; p = 0.80). CSE-exposed tissues demonstrated a dose-dependent decrease in Bcl2 expression and increases in active caspase 3 and nuclear fragmentation in both amnion and chorion cells compared to controls. In summary, fetal membranes exposed to CSE manifest evidence of OS and apoptosis. The differential pattern of prostanoid production observed in this study supports the hypothesis that an alternate non-inflammatory pathway mediated by OS and apoptosis in pPROM may promote proteolysis resulting in membrane weakening and rupture.
authors.affiliation: Department of Epidemiology, Rollins School of Public Health, Emory University, 1518 Clifton Ave NE, Atlanta, GA 30322, USA. rmenon3@emory.edu
authors.forename: R
authors.initials: R
authors.lastname: Menon
authors.forename: S J
authors.initials: SJ
authors.lastname: Fortunato
authors.forename: J
authors.initials: J
authors.lastname: Yu
authors.forename: G L
authors.initials: GL
authors.lastname: Milne
authors.forename: S
authors.initials: S
authors.lastname: Sanchez
authors.forename: C O
authors.initials: CO
authors.lastname: Drobek
authors.forename: M
authors.initials: M
authors.lastname: Lappas
authors.forename: R N
authors.initials: RN
authors.lastname: Taylor
mesh_terms: D017209:Apoptosis / Q000187:drug effects*; D053148:Caspase 3 / Q000096:biosynthesis; D015237:Dinoprost / Q000096:biosynthesis; D005321:Extraembryonic Membranes / Q000187:drug effects* / Q000473:pathology; D028441:F2-Isoprostanes / Q000096:biosynthesis; D005260:Female; D005322:Premature Rupture of Fetal Membranes / Q000139:chemically induced*; D006801:Humans; D018384:Oxidative Stress / Q000187:drug effects*; D011247:Pregnancy; D012906:Smoke / Q000009:adverse effects*; D012907:Smoking / Q000009:adverse effects*; D051025:bcl-Associated Death Protein / Q000096:biosynthesis
chemical_list: D028441:F2-Isoprostanes; D012906:Smoke; D051025:bcl-Associated Death Protein; D015237:Dinoprost; D053148:Caspase 3